The present invention relates generally to mass spectrometry, and more particularly to solvent-free matrix deposition apparatus and methods using sublimation for imaging mass spectrometry.
Matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI-IMS) is a revolutionary and emerging technique for profiling and mapping biomolecules and studying related biological processes directly from the samples. MALDI-IMS is discussed by M. Stoeckli, et al., in “Imaging mass spectrometry: A new technology for the analysis of protein expression in mammalian tissues”, Nat Med, 7, 2001, 493-496, for example.
Based on its potential of simultaneously providing identification and spatial localization of most molecules of interest in complicated biological samples such as tissues and cells, imaging mass spectrometry has been recognized as a promising tool for numerous applications including peptide/protein, lipid, metabolite analysis, biomarker discovery, drug biodistribution monitoring, molecular mechanism investigation, etc. This is discussed by R. Lemair, et al., in “MALDI-MS Direct Tissue. Analysis of Proteins: Improving Signal Sensitivity Using Organic Treatments”, Anal Chem, 78, 2006, 7145-7153, for example.
Uniformly coating the matrix onto sample surfaces is one of the most critical experimental factors in MALDI-IMS. The reproducibility and quality of MALDI imaging analyses are greatly affected by the size and homogeneity of the matrix cluster/crystal as well as the inevitable analyte spreading caused by solvents used for matrix deposition.
Currently, there are two generally-used methods for matrix deposition: spraying and spotting. Both of them have a common problem of using solvents as media to transfer the matrix onto the sample surface. It is impossible to avoid lateral movement of the analytes with the existence of solvents, which reduces the signal intensity and resolution of molecular imaging. In addition, spraying and spotting techniques have other limitations, such as high irreproducibility, poor control of matrix/analyte incorporation, slow coating process, and alkali metal contamination (Nat or K).
A number of U.S. patents have been issued relating to ionization mass spectrometry. U.S. Pat. No. 5,643,800 entitled “Method of preparing a sample for analysis by laser desorption ionization mass spectrometry” issued Jul. 1, 1997, U.S. Pat. No. 5,854,486 entitled “Method and apparatus for MALDI mass spectrometry” issued Dec. 29, 1998, and U.S. Pat. No. 6,104,028 entitled “Volatile matrices for matrix-assisted laser desorption/ionization mass spectrometry” issued Aug. 15, 2000.
As evidenced by its abstract, U.S. Pat. No. 5,643,800 discloses “a sample preparation system and method that can be used with all types of analyte materials, that produces homogeneously deposited crystals across a sample surface, and that lends itself to automation. In this system and method, analyte crystallization is caused by lyophilization. A homogeneous analyte/solvent mixture is placed on a sample surface. The mixture is frozen, then the solvent is sublimated through the application of a vacuum. A homogenous distribution of analyte crystals across the sample surface results.” Thus, U.S. Pat. No. 5,643,800 discloses sublimation of a solvent that has been placed on a surface to create a crystalline surface on a sample.
As evidenced by its abstract, U.S. Pat. No. 5,854,486 discloses that a “thin uniform film of matrix material is deposited from the gas phase on to a substrate for use in Matrix Assisted Laser Desorption and Ionization (MALDI) spectroscopy. The thin uniform film of material may be overcoated with another film of material which has a higher vapor pressure than the matrix material to prevent the matrix material from evaporating during storage and during substantial time in the vacuum environment of the mass spectrometer.” The Summary of the Invention section of U.S. Pat. No. 5,854,486 states that “A thin, uniform film of matrix material is deposited on a substrate by sublimation of the solid matrix material or by other vapor deposition methods.” U.S. Pat. No. 5,854,486 discloses a column 5, lines 20-25 that “The analyte material 50 may be deposited on the matrix material of layer 20 by a number of techniques. The easiest of such techniques is merely to drop a drop of solution containing the analyte material, a volatile solvent, and optionally matrix material on the surface of the layer 20 and to allow the volatile component of the solution to evaporate.”
The abstract of U.S. Pat. No. 6,104,028 states that a “sample preparation method is disclosed for volatilization and mass spectrometric analysis of nonvolatile high molecular weight molecules. Photoabsorbing molecules having significant sublimation rates at room temperature under vacuum, and preferably containing hydroxy functionalities, are disclosed for use as matrices in matrix-assisted laser desorption/ionization mass spectrometry. The samples are typically cooled in the mass spectrometer to temperatures significantly below room temperature.” The Summary of the Invention section of U.S. Pat. No. 6,104,028 discloses “a method for determining the mass of a large organic molecule” and that the sample preparation method uses “liquids or low sublimation temperature solids as matrices because such systems generally enable lower desorption/ionization temperatures.” However, U.S. Pat. No. 6,104,028 discloses that “In creating the matrix:molecule mixture, for example, by dissolving the large organic molecule in a solution containing the matrix, one of skill in the art will understand that the solution containing the matrix may generally contain one or more solvents.”
However, none of the above-cited patents discloses or suggests formation of a solvent-free matrix using sublimation techniques. In addition, there are no conventional molecular matrix deposition systems that have matrix (source) and sample (target) movement systems or stages for optimizing sublimation-condensation position and mixed matrix deposition. Furthermore, no conventional molecular matrix deposition system employs apparatus that permits substantially homogeneous deposition of molecules from multiple matrices on one or multiple samples. Finally, no conventional molecular matrix deposition system employs apparatus that permits deposition of matrix molecules on multiple samples at substantially the same time.
It would be desirable to overcome the limitations offered by conventional matrix deposition approaches. It would be desirable to have solvent-free matrix deposition apparatus and methods for use in imaging mass spectrometry and other surface coating processes. It would be desirable to have apparatus and methods that provide uniform gas-phase molecular solvent-free matrix deposition for imaging mass spectrometry and other surface coating processes. It would be desirable to have apparatus and methods that form a solvent-free matrix on a sample using sublimation techniques. It would be desirable to have apparatus and methods that advantageously employ uniform gas-phase molecular solvent-free matrix deposition to coat surfaces of materials.